GABA-modulin is an endogenous synaptic membrane protein of 17,000 MW which inhibits the binding of 3H-GABA and the GABA-induced stimulation of 3H-diazepam binding to brain synaptic membranes. GABA-modulin is a substrate for cAMP-dependent and Ca2+-dependent protein kinases. Different sites in GABA-modulin are phosphorylated by the two classes of protein kinases. Extensive phosphorylation of GABA-modulin by the cAMP-dependent protein kinase results in the lost of its inhibitory effects on GABA binding, whereas Ca2+/calmodulin-dependent phosphorylation has no effect on its biological activity. A GABA-modulin-like protein is present in membranes prepared from bovine adrenal medulla and adrenal chromaffin cells maintained in primary culture as revealed by radioimmunoassay. Endogenous protein phosphorylation stimulated by Ca2+, Ca2+ plus phospholipid, and cAMP in adrenal medullary membranes reveals the presence of a phosphoprotein of similar molecular weight as GABA-modulin. Similarly, a phosphorylated GABA-modulin-like protein is immunoprecipitated from acid extracts of 32P-labeled chromaffin cells. Therefore, the bovine adrenal chromaffin cell may be a suitable model in which to study the regulation of GABA-modulin phosphorylation and its functional role in the GABA/benzodiazepine receptor complex.